Name: rat heterotopic abdominal heart/single-lung transplantation in a volume-loaded configuration
Uploaded: 2015-05-29T13:00:00.000+00:00
Duration: 6 min 16 s
Description: Watch this Scientific Journal Video about Rat Heterotopic Abdominal Heart/Single-lung Transplantation in a Volume-loaded Configuration at JoVE.com

Mark J. Kearns (Doktorlar ve Kanada Royal College of Surgeons aracılığıyla verilen) UBC Klinisyen Araştırmacı Programı ve UBC 4YF Doktora Bursu destek alır. John H. Boyd Sağlığı Araştırma Scholar için Ulusal Sanatoryum Derneği ve Michael Smith Vakfı. CIHR aracılığıyla Fon. Yazarlar kurulum ve perfüzyon ekipmanın bakım ile yardım için Dr. M. Allard ve Richard Wamboldt teşekkür etmek istiyorum.

Tüm hayvanların barındırıldığı ve Ulusal ve bakım ve laboratuar hayvanlarının kullanımı için kurumsal kurallar ile uyumlu bakım için. Bu protokol Etik onay British Columbia Hayvan Bakım Komitesi Üniversitesi tarafından verildi. 300 arasındaki ağırlığındaki erkek Sprague-Dawley sıçanları - 450 g bu protokol için kullanılmıştır. 1. Donör Operasyonu <ol><li> 3-yollu uzun intravenöz (IV), kateter tüp bağlanan bir silindirik bir şişeye kardiyopleji (RT), yaklaşık olarak 100 ml yazıldı. Yerçekimi tarafından kardiyopleji teslim edilmesini sağlayarak, yaklaşık 80cm operatif yüzeye yukarıda şişeyi yükseltmesine bir stand kullanın. </li><li> Yeterince yapılarını görselleştirmek için, cerrahi loupes bir çift veya bir diseksiyon mikroskobu birini kullanın. NOT: Şu anda 3.4 ile binoküler mikroskop çalışma kullanın - 21.3X büyütme. </li><li> Bir anestezik odasında donör koyun ve 4 ile anestezi neden -% 5 izofluran. </li&gt; <li> Bir işletim platformuna sıçan aktarın ve 1 ile burun konisi ile anestezi korumak -% 2 izofluran. Kuruluğunu önlemek için hayvanın gözlerine veteriner merhem sürün. 25 G iğne ile midazolam (2 mg / kg) intraperitoneal olarak yönetmek. </li><li> Cerrahi makası kullanarak, alt çeneye xiphisternum gelen donör tıraş. (Sonradan IV erişimi için), sol kasık küçük bir bölgesini tıraş. Operatif yüzeylere bir Tüyleri ajanı uygulayın, yaklaşık 5 dakika bekleyin ve gazlı bez parçası ile saç kaldırmak. </li><li> Bir povidon-iyot veya chlorexidine tabanlı bir çözüm olan cerrahi siteleri (biz sadece chlorhexidine kullanın) hazırla. % 0.5 lidokain subkutan - 0.1 ile insizyonel siteleri sızmak. </li><li> Anestezi derinliği ve vital bulguların izlenmesi için ücretsiz sağ hindlimb bırakarak, yapışkan bant ile çalışma platformu ön ayakları ve sol hindlimb sabitleyin. </li><li> Pedal tutam uygun anestezi derinliği olduktan sonra kullanarak, sol kasık kırışık bir kesi yapmak22-bıçak neşter. Künt diseksiyon ile, sol ana femoral ven ortaya çıkarmak ve şöyle IV erişim elde: <ol><li> Yavaşça femoral ven örten doku geri ve 24 G IV damar cannulate. Tuzlu dolu IV boru kısa bir uzunluğa IV bağlayın ve yapışkan bant ile yerine tüp sabitleyin. NOT: sıçanlarda femoral venöz kanülasyon için prosedür Jespersen ve arkadaşları 14 tarafından başka açıklanmıştır. </li><li> IV doğru konumlandırma sağlamak için kan aspire hafifçe IV boru bir tuzlu dolu 10 ml şırınga bağlayın ve. </li><li> IV tüp aracılığıyla fraksiyone olmayan heparine 500 IU, ve daha sonra 3 ile tüp yıkayın - - 300 enjekte serum fizyolojik 5 ml. </li></ol></li><li> Bunun ardından, verici tracheotomize: <ol><li> 22-bıçaklı neşter kullanılarak juguler çentik ve mandibulanın arasındaki yumuşak dokularda bir orta hat kesi olun. Metzen kullanarak orta hatta tiroid bezinin kapsülü NüfuzKünt diseksiyon kullanarak baum makas, ve ayrı onun loblar. </li><li> Künt diseksiyon kullanarak, trakea ön yüzeyini açığa orta hatta boyun askısı kasları ayırın. </li><li> Açık açık trakea etrafında bir çember uçağı incelemek için kavisli bir forseps kullanın. 4-0 ipek kravat ile trakea kuşatmak. </li><li> Iris makas kullanarak, tiroid kıkırdak yaklaşık 5mm aşağı, ön trakea içinde enine bir kesi yapmak. Yavaşça trakeal kanül (14 G IV) tanıtmak ve 4-0 ipek kravat kullanarak yerine sabitleyin. </li><li> Mekanik ventilatöre trakeal kanül bağlayın. Ventilatör devresi aracılığıyla oksijen ve izofluran akışını Redirect ve hızı ve ağırlığı 13 tarafından tahmin tidal hacimde donör havalandırın. </li></ol></li><li> Xipisternum altına juguler çentik at kesi uzanan, (22 bıçaklı neşter kullanılarak) göğüs orta hatta bir kesi yapmak. </li><li> Orta hatta kalıyorumBir kemik kesici kullanılarak medyan sternotomi gerçekleştirin. Kendinden istinat retraktörün ile sternum kenarlarını geri çekin. Perikard ve plevra boşlukları girin. </li><li> Bir timektomi gerçekleştirin. İlk açık açık orta hatta timus bölmek ve sonra künt ve keskin diseksiyon bir arada kullanarak çevre yapılardan ayırmak için en kolay yoldur. NOT: timüs uzakta üstün sternum kenarlardan diseksiyon sırasında internal torasik arterlerin kökeni yaralı olabilir. Kanamayı önlemek için, hemostatik klipler bu noktalarda timus çıkarmadan önce tatbik edilebilir. </li><li> Metzenbaum makas kullanarak ve / veya keskin Lauer, inferior vena kava (IVC) uzak yağ peri-kaval teşrih. IVC altta diyaframa, superior cavo-atriyal kavşağına yağ ve bağ dokusu görece özgür olduğundan emin olun. </li><li> Keskin Lauer, çevresel ücretsiz superior vena kava (SVK) kullanma ve 4-0 ipek kravat ile kuşatmak. </li><li> Sonraki, teşrihçevredeki yapılardan ücretsiz vena kava sol ve proksimal ve distalde 4-0 ipek kravatlar ile bağlanır. Sol subklavian arteri ortaya çıkarmak için cava müdahale kısmını rezeke. </li><li> Çevresel ücretsiz keskin Lauer kullanarak arkus aorta gemiler. Innominate artere yakın ve uzak cerrahi klipleri uygulayın ve kliplerin arasına bölün. Sol karotis arter ve sol subklavyen arter un-kırpılmış bırakın. </li><li> Sonra, 24 G IV kateter ile SVC cannulate. Önceden yerleştirilen 4-0 ipek kravat ile kateter sabitleyin. </li><li> Bir çare, ama nazik greft hasat için hazırlanın. </li><li> Keskin bir makas kullanarak, diyafragma yüzeyine IVC sadece üstün bölün. Yakındaki havayolu yapıları yaralı olmadığını sağlanması, onun kanülasyon sitesine SVC üstün bölün. </li><li> Vantilatör kapatın ve trakeal kanül ayırın. Proksimale trakea transect. </li><li> Forseps ile kesilen trakea tutun ve kalp kaldırmakve akciğerler en blok. Kalp-akciğer greft kaldırılır gibi bu keskin diseksiyon ile nazik çekiş gerektirecektir. Altta yatan özofagus trakea ayırın. Bunun büyük bir kısmı hasattan sonra bozulmadan kalır, böylece inen aort yaralanmasına kaçının. </li></ol> Kalp-akciğer Greftinin 2. Hazırlık NOT: Prosedürün bu bölümünü tamamlarken, bir asistan alıcı kemirgen anestezi ve greft implantasyonu için hazırlık olmalıdır. <ol><li> Aşağı bakacak şekilde kalbin ön yüzeyi ile cerrahi gazlı bez üzerine kalp-akciğer greft yerleştirin. Keskin diseksiyon kullanarak, arka hava yollarına yaralanmasına kaçınarak, herhangi bir kalıntı özofagus çıkarın. </li><li> Inen aorta bulun ve bir 16 G künt uçlu kanül yerleştirin. 4-0 ipek kravat ile yerine kanül sabitleyin. </li><li> Aortik ark damarları bulun ve sol subklavian arter cerrahi klip uygulayın. Un-kırpılmış sol karotis arter bırakın alt içinizleyen de-havalandırma. </li><li> 16 G aort kanül Kardiyoplejisi boru bağlayarak kardiyoplejisinin ilk dozunu. Karotis arter geçici kardiyoplejisinin yeterli teslim edilmesini sağlamak için forseps ile tıkanmıştır gerekir. Not: bağlı aortik kanül sahip bir yararı arzu edildiği gibi kardiyopleji, sürekli ve / veya kesintili olarak tatbik edilebilir olmasıdır. 45 sn - 30 5 ml bolus 15 dk - aralıklı doz için, genellikle kardiyoplejisinin her 10 teslim. </li><li> Metzenbaum makas kullanarak ve / veya keskin Lauer, uzak aortadan peri-aort yağ teşrih. Aort kendi kanulasyon sola subklavian arter (distal kemer) den yağ ve bağ dokusu görece özgür olduğundan emin olun. </li><li> Sonra, keskin bir Lauer kullanarak sol ana bronş maruz ve 4-0 ipek kravat proksimale Arter. Iris makası kullanarak dikilen sola bronş distalinden transect. Sol pulmoner bağlanarak bir sol pnömonektomi gerçekleştirin4-0 ipek kravatlar ile arter ve ven. Ligature distalinde transect ve sol akciğeri çıkarın. </li><li> Yukarıdaki gibi, sağ üst ve orta lob hariç diğer tüm akciğer lobları kaldırın. Hastaya lobektomi gerçekleştirilmesi sürecinde IVC yaralanmasına kaçının. </li><li> Aort kanül Kardiyoplejisi boru bağlayın ve alıcı hazırlanan edilirken sürekli kardiyoplejisine yönetmek. Steril bir kaba (örneğin, bir bardak) kalp-akciğer greft yerleştirin. </li></ol> 3. Alıcı Operasyonu <ol><li> Yukarıda, donör operasyonu başına alıcı anestezisi. Kuruluk gelen hayvanın gözleri korumak için veteriner merhem kullanın. Pedal tutam tarafından sıkça anestezi onaylayın. </li><li> Vital bulguları ve anestezi derinliği izlemek için ücretsiz sağ ön ayakları bırakarak, donör operasyonu başına bu kez hayvan yerleştirin. </li><li> Xiphisternum gelen penise karın Tıraş. (Sonradan IV erişimi için), sol kasık küçük bir bölgesini tıraş. Uygula biroperatif yüzeylere ajan tüylerini giderildikten, yaklaşık 5 dakika bekleyin ve gazlı bez parçası ile saç kaldırmak. </li><li> Bir povidon-iyot veya chlorexidine tabanlı bir çözüm olan cerrahi siteleri hazırlayın. % 0.5 lidokain subkutan - 0.1 ile insizyonel siteleri sızmak. </li><li> Tracheotomize ve yukarıda belirtildiği şekilde alıcı havalandırın. Alternatif olarak, burun konisi anestezi altında alıcı korumak. </li><li> Yukarıda tarif edildiği gibi femoral damara bir kateter tanıtılması. IV tüp aracılığıyla fraksiyone olmayan heparine 500 IU, ve daha sonra 3 ile tüp yıkayın - - 300 enjekte serum fizyolojik 5 ml. </li><li> Penise xiphisternum bir 22-bıçak neşter ile orta hat karın kesi yaparak bir laparotomi gerçekleştirin. Kendi kendine istinat Retraktörü kullanarak karın duvarı çekin. Sonra, superior ve sıçan sol tarafa bağırsakları geri çekin. Sıcak, tuzlu batırılmış gazlı bez onları sarın. </li><li> Keskin örten yeniden aracılığıyla diseksiyon ile VKİ ve abdominal aorta Açığatro-periton yağ. </li><li> Kavisli damar kelepçe mevcut ve hazır bulundurun. </li><li> Çevresel proksimale ve distale IVC ve aort özgür ve 4-0 ipek kravatlar ile kuşatmak. Bu siteler arasında boşluk 3 cm - yaklaşık 2 olmalıdır. </li><li> Dikkatle IVC ve aort hem yeterli bir bölümünün kelepçe çene yukansında yer alır sağlanması, eğri damar kelepçesini uygulanır. </li><li> Bir tuzlu dolu 1 ml şırınga bağlı 25 G iğne ile IVC ön duvarında bir kesi yapmak. Donör IVC deliğin uzunluğu maç için Potts makas ile kesi uzatın. </li><li> Kendi kabından kalp-akciğer greft çıkarın ve kardiyoplejisinin bağlantısını kesmek. Venöz anastomoz gerçekleştirmek için en uygun pozisyonda alıcının karın greft yerleştirin. NOT: kalp-akciğer greft sonuçta karın yıllardan doğru apeks işaret ile, hafifçe eğik odaklı olacak alt kadranda bıraktı. </li><li> Topuk Güvenlive ayak 9-0 naylon sütür ile anastomoz biter. Dikiş uzun koluna bağlı iğne, daha sonra bağlı olan sütür kısa bir ucunu bırakarak, her iki ucunda güvenli bir düğüm. Kardiyoplejisinin bir doz yönetme. </li><li> Moda çalışan olarak, karşıt kısa dikiş koluna dikiş hattı ve kravat tam bir buçuk. Kardiyoplejisinin bir doz yönetme. </li><li> Sütür hattının diğer yarısını tamamlayın ve aşağı kravat. Venöz anastomoz tamamlandı. Kardiyopleji bir dozunun tatbik edilmesi </li><li> Orient karın yıllardan karşı kalbin apeks işaret ile kalp-akciğer greft alt kadranda bıraktı. Venöz anastomoz kıvrılmış veya bükülmüş olmadığından emin olun. </li><li> Gerilimsiz ve un-Dirsekli konfigürasyonlarda IVC ve aort anastomoz hem bakımını alıcı aorta ulaşmak için gerekli olacak donör aort uzunluğunu değerlendirin. </li><li> Bir iris makas ile (sol subklavian arter distalinde) inen aorta bölün. </li><li> Bir incisio Yapn, bir tuzlu su ile doldurulmuş 1 ml şırınga bağlı bir 25 G iğne ile alıcı aort ön duvarında. Donör aort deliğinin uzunluğu maç için Potts makas ile kesi uzatın. </li><li> 9-0 naylon dikiş kullanılarak aort alıcıya donör aort burnunu (üstün yönü) Çapa. Ardından, aortun medial iğne geçmek ve moda çalışan anastomoz yarısını tamamlamak. </li><li> Aort anastomoz topuk, sütür hattının (dikiş yönünde bir geçiş) yönünü ters ve moda çalışan anastomoz lateral yarısını tamamlamak. </li><li> Yavaşça SVC IV kateter kaldırmak ve cerrahi klip ile SVC Arter. </li><li> De-hava un kırpılmış sol karotis arter bularak aorta. Havanın serbestçe tahliye sağlayacak bir dik bir konumda arteri açık tutmak. </li><li> Kısaca d kavisli damar kelepçe çeneleri açılır ve (karotid arter kanamaya izinE-hava) 2 - 3 sn. Damar kelepçesini yeniden uygulayın. </li><li> Karotid arter cerrahi bir klip uygulayın. Kavisli damar kelepçesini çıkarın. </li><li> Sütür hatlarında kanama olup olmadığını kontrol edin. Varsa, 9-0 naylon sütür (kanamanın şiddetine bağlı olarak) uzunluğunda bir cerrahi gazlı bez ya da tamir nazik sıkıştırma uygulayın. </li><li> Donör kalp dakika içinde yenerek devam etmesi gerekir. </li><li> Alıcının vital bulgular, anestezi derinliği yakın ilgi ve gerektiği gibi ses canlandırma yönetmek. </li><li> Ya deneysel uç noktaları bağlı hayvanın karın kapatın ve anestezi kurtarmak izin veya kardiyovasküler değerlendirme için hayvan hazırlar. NOT: greft değerlendirme örnekleri yük-bağımlı ve yük bağımsız hemodinami in vivo tedbirler, in vitro Langendorff&#39;a işlevin önlemleri ve çalışma kalp modları ve (hayatta kalma cerrahi) ekokardiyografik veya MR incelemeleri dahil. </li><li> AtAkut Terminal deneyler sonunda, hayvanlar kan kaybından ötenazi. </li></ol>

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Yazarlar ifşa hiçbir şey yok.

Burada anlatılan teknikle Başarı çeşitli faktörlere esas alınacaktır. Aralarında Key üniforma greft soğutma ile tam kardioplejik tutuklama, toplam iskemik süresini en aza indirmek sağlanması, güvenli ve minimal kan kaybı ile ilişkili olan titiz operatif teknik benimseyen, hem donör ve alıcı hayvanların istikrarın sağlanması ve yeterli de-havalandırma greft olacaktır. Yukarıda kabul gibi, tekniğin teknik karmaşıklığı onun baş sınırlamadır. Biz birçok bak?...

The first rodent model of heterotopic abdominal heart transplantation (HAHT) was described by Abbott and colleagues in 19641. This technique, and subsequent modifications have been widely applied to characterize transplant graft function and immunologic status. The majority of HAHT techniques described involve a non-volume loaded heart2,3. Models of HAHT involving volume-loaded ventricles have been described, but they are frequently limited in one or more respects.
Heterotopic abdominal heart-lung transplantation (HAHLT) with a volume-loaded left ventricle (LV) has been described previously. Chen and colleagues4, and subsequently Ibrahim and colleagues5 described HAHLT with a single aorto-aortic (donor ascending to recipient abdominal aorta) anastomosis. The only volume load presented to the ventricle in this circulation is the coronary venous return. Asfour and colleagues described a HAHT technique in which the lung circuit was eliminated by anastomosing donor pulmonary artery (PA) to donor left atrium (LA)6. In this circulation, venous inflow to right ventricle (RV) occurs via a donor SVC to recipient IVC anastomosis, and the subsequent LV load is ejected into the aorto-aortic anastomosis. Cardiac function was partially assessed in vivo, and also in vitro using a Langendorff rig. Figueiredo and colleagues described a HAHLT model similar to our own7, but in mice. Venous inflow to the RV occurs via donor SVC to recipient IVC anastomosis. Blood subsequently passes through the single lung circulation and LV load is ejected into the aorto-aortic anastomosis. Cardiac function in their study was assessed by magnetic resonance imaging (MRI). Wen and colleagues described a unique HAHT technique in which the LV is loaded by means of a recipient aorta to donor LA anastomosis8. The LV, therefore, fills at systemic pressures. Cardiac function, and whether LV stroke volume is ejected antegradely in their model was not assessed.
Many of the techniques referenced above involve non-physiologic LV loading conditions, including the techniques whose partial LV load is represented only by coronary venous return. On the other hand, many techniques do approach physiologic LV loading. The majority of these techniques, as with the technique of Asfour and colleagues, omit the pulmonary circulation and utilize a donor PA to donor LA anastomosis6,9. The circulation described by Galinanes and colleagues10 employs a direct recipient cava to donor LA anastomosis, omitting the pulmonary circulation and the right heart. Yokoyama and colleagues achieve the same effect by ligating the donor PA and creating an interatrial communication in the donor heart (omitting donor lung and right heart circulations)11. The circulation of Maruyama and colleagues12 involves an anastomosis between donor left PA and recipient Ao, which permits LV filling via the pulmonary circulation as a conduit, but effectively excludes the right heart.
In cases where near physiologic loading conditions were met, we advance the technique of HAHLT in 2 major respects. First, to our knowledge, the exact configuration we report has not been described in rats. It is possibly the most versatile circulation for investigators wishing to study the physiology, structure, and immunology of the transplanted heart-lung graft. Second, we describe how the function of the transplant graft can be directly characterized in vivo. For this application, pressure-volume conductance catheters can be introduced directly into the LV apex of the transplant graft, which allows for complete cardiac functional characterization. 
The technique described here can be applied to both acute and chronic studies of transplant graft function, while the functional assessment may be performed either in vivo or in vitro. We present a model in which the loading conditions can be near physiologic, however the degree of ventricular loading may be manipulated both acutely and chronically by diverting venous return towards or away from the graft. Afterload conditions can also be manipulated. Because the lung and its airway are retained in this transplant configuration, investigators can re-ventilate the donor lung acutely. Uniquely, lung re-ventilation changes the composition of blood perfusing the transplant coronary arteries. Under non-ventilated conditions, blood ejected from the donor aorta is deoxygenated, and mixes with oxygenated blood in the recipient aorta. Under acutely ventilated conditions, ejected blood becomes oxygenated. Thus, transplant graft function can be compared under ventilated and non-ventilated conditions, and also under variably loaded conditions.
The protocol below describes important modifications to previously described HAHLT donor and recipient operations. It also describes an optimal technique for protecting the transplant graft throughout the period of ischemia (time between donor explant and recipient implant). Advantages of this technique include physiologic conditions potentially approaching that of an orthotopically transplanted graft, and a wide range of investigative applications. An important limitation is its technical complexity. With adequate mentoring and practice, the advantages of this technique will likely outweigh the challenges in adopting it.

<table><tbody><tr><td>Celsior Cardioplegic Soution</td><td>Genzyme</td><td></td><td>The solution is kept on ice throughout the procedure. We prepare our own solution, with slight modifications.</td></tr><tr><td>Rodent Ventilator</td><td>Harvard Apparatus</td><td>Model 683</td><td></td></tr><tr><td>Vital Sign Monitor</td><td>Nonin</td><td>Model 9847V</td><td>Displays SpO&lt;sub&gt;2&lt;/sub&gt; and heart rate.</td></tr><tr><td>IV Cannulae</td><td>Jelco</td><td>3063</td><td>24 - 26 G x 3/4&quot; cannulae.</td></tr><tr><td>IV Tubing</td><td>CareFusion</td><td>MP9259-C</td><td>Short-length connector tubing (18 cm).</td></tr><tr><td>Surgical Clips</td><td>Teleflex Medical</td><td>001204</td><td>Horizon titanium ligating clips.</td></tr><tr><td>Sutures</td><td>Ethicon, Sharpoint</td><td>LA54G, AK-0107</td><td>3-0 silk reel, and 9-0 prolene suture (single-armed, DR5 needle).</td></tr><tr><td>Surgical Instruments</td><td>Not Applicable</td><td>Not Applicable</td><td>The instruments used are generic, and can be purchased from any surgical supply company.</td></tr></tbody></table>

rat heterotopic abdominal heart/single-lung transplantation in a volume-loaded configuration

Herein, we describe a novel technique for heterotopic abdominal heart-lung transplantation (HAHLT) in rats. The configuration of the transplant graft involves anastomosis of donor inferior vena cava (IVC) to recipient IVC, and donor ascending aorta (Ao) to recipient abdominal Ao. The right upper and middle lung lobes are preserved and function as conduits for blood flow from right heart to left heart.
There are several advantages to using this technique, and it lends itself to a broad range of applications. Because the graft is transplanted in a configuration that allows for dyamic volume-loading, cardiac function may be directly assessed in vivo. The use of pressure-volume conductance catheters permits characterization of load-dependent and load-independent hemodynamic parameters. The graft may be converted to a loaded configuration by applying a clamp to the recipient&#8217;s infra-hepatic IVC. We describe modified surgical techniques for both donor and recipient operations, and an ideal myocardial protection strategy. Depending on the experimental aim, this model may be adapted for use in both acute and chronic studies of graft function, immunologic status, and variable ventricular loading conditions. The conducting airways to the transplanted lung are preserved, and allow for acute lung re-ventilation. This facilitates analysis of the effects of the mixed venous and arterial blood providing coronary perfusion to the graft.
A limitation of this model is its technical complexity. There is a significant learning curve for new operators, who should ideally be mentored in the technique. A surgical training background is advantageous for those wishing to apply this model. Despite its complexity, we aim to present the model in a clear and easily applicable format. Because of the physiologic similarity of this model to orthotopic transplantation, and its broad range of study applications, the effort invested in learning the technique is likely to be worthwhile.

Yukarıda tarif edilen HAHLT tekniği son derece teknik ve ayrıntı. Tablo yakın ilgi gerektiren 1 golleri başarısız prosedürlere karşı başarılı ile ilişkili önemli faktörlerden bazıları ve teknik zorlukları gidermek için bir rehber olarak kullanılabilir. Alıcı aort kelepçelenmemiş sonra, greft koroner arterler oksijenli kan ile doldurmak için görülmelidir. Buna göre, miyokard pembe ve iyi perfüze haline gelmelidir. Teknik olarak ...

Watch this Scientific Journal Video about Rat Heterotopic Abdominal Heart/Single-lung Transplantation in a Volume-loaded Configuration at JoVE.com

Rat Heterotopic Abdominal Heart/Single-lung Transplantation in a Volume-loaded Configuration

We describe a novel technique for heterotopic abdominal heart-lung transplantation (HAHLT) in rats. The transplant configuration results in a partially loaded graft circulation, allowing direct functional assessment. This model may be employed for acute or chronic studies of function and immunologic status of the transplanted graft.

Sıçan Heterotopik Abdominal Kalp / Bir Ses yüklenen Yapılandırması Tek akciğer transplantasyonu

Herein, we describe a novel technique for heterotopic abdominal heart-lung transplantation (HAHLT) in rats. The configuration of the transplant graft ...

rat-heterotopic-abdominal-heartsingle-lung-transplantation-volume

Research

JoVE Journal

Medicine

8.3K Görüntüleme.  UBC James Hogg Research Center, St. Paul's Hospital. We describe a novel technique for heterotopic abdominal heart-lung transplantation (HAHLT) in rats. The transplant configuration results in a partially loaded graft circulation, allowing direct functional assessment. This model may be employed for acute or chronic studies of function and immunologic status of the transplanted graft.

Video: Rat Heterotopic Abdominal Heart/Single-lung Transplantation in a Volume-loaded Configuration

Heterotopic Heart Transplantation in Mice

The mouse heterotopic heart transplantation has been used widely since it was introduced by Drs. Corry and Russell in 1973. It is particularly valuable for studying rejection and immune response now that newer transgenic and gene knockout mice are available, and a large number of immunologic reagents have been developed. The heart transplant model is less stringent than the skin transplant models, although technically more challenging. We have developed a modified technique and have completed over 1000 successful cases of heterotopic heart transplantation in mice. When making anastomosis of the ascending aorta and abdominal aorta, two stay sutures are placed at the proximal and distal apexes of recipient abdominal aorta with the donor s ascending aorta, then using 11-0 suture for anastomosis on both side of aorta with continuing sutures. The stay sutures make the anastomosis easier and 11-0 is an ideal suture size to avoid bleeding and thrombosis.
When making anastomosis of pulmonary artery and inferior vena cava, two stay sutures are made at the proximal apex and distal apex of the recipient s inferior vena cava with the donor s pulmonary artery. The left wall of the inferior vena cava and donor s pulmonary artery is closed with continuing sutures in the inside of the inferior vena cava after, one knot with the proximal apex stay suture the right wall of the inferior vena cava and the donor s pulmonary artery are closed with continuing sutures outside the inferior vena cave with 10-0 sutures. This method is easier to perform because anastomosis is made just on the one side of the inferior vena cava and 10-0 sutures is the right size to avoid bleeding and thrombosis. In this article, we provide details of the technique to supplement the video.

The mouse heterotopic heart transplantation model has been proven by many investigators to be an important method for studying mechanisms of rejection and immune response. However, the techniques involved are still challenging. By modifying standard techniques we have had success with more than 1000 transplants.

Farelerde Heterotopik Kalp Nakli

The mouse heterotopic heart transplantation has been used widely since it was introduced by Drs. Corry and Russell in 1973. It is particularly valuable ...

Biyoloji

Heterotopic Auxiliary Rat Liver Transplantation With Flow-regulated Portal Vein Arterialization in Acute Hepatic Failure

In acute hepatic failure auxiliary liver transplantation is an interesting alternative approach. The aim is to provide a temporary support until the failing native liver has regenerated.1-3 The APOLT-method, the orthotopic implantation of auxiliary segments- averts most of the technical problems. However this method necessitates extensive resections of both the native liver and the graft.4 In 1998, Erhard developed the heterotopic auxiliary liver transplantation (HALT) utilizing portal vein arterialization (PVA) (Figure 1). This technique showed promising initial clinical results.5-6 We developed a HALT-technique with flow-regulated PVA in the rat to examine the influence of flow-regulated PVA on graft morphology and function (Figure 2).
A liver graft reduced to 30 % of its original size, was heterotopically implanted in the right renal region of the recipient after explantation of the right kidney.&#160; The infra-hepatic caval vein of the graft was anastomosed with the infrahepatic caval vein of the recipient. The arterialization of the donor&#8217;s portal vein was carried out via the recipient&#8217;s right renal artery with the stent technique. The blood-flow regulation of the arterialized portal vein was achieved with the use of a stent with an internal diameter of 0.3 mm. The celiac trunk of the graft was end-to-side anastomosed with the recipient&#8217;s aorta and the bile duct was implanted into the duodenum. A subtotal resection of the native liver was performed to induce acute hepatic failure. 7
In this manner 112 transplantations were performed. The perioperative survival rate was 90% and the 6-week survival rate was 80%. Six weeks after operation, the native liver regenerated, showing an increase in weight from 2.3&#177;0.8 g to 9.8&#177;1 g. At this time, the graft&#8217;s weight decreased from 3.3&#177;0.8 g to 2.3&#177;0.8 g.
We were able to obtain promising long-term results in terms of graft morphology and function. HALT with flow-regulated PVA reliably bridges acute hepatic failure until the native liver regenerates.

Auxiliary liver transplantation provides a temporary support in acute hepatic failure, until regeneration of the failing liver. The heterotopic auxiliary liver transplantation (HALT) with portal vein arterialization (PVA) renders sufficient liver function. We developed an analogous technique in the rat, to examine the influence of the portal vein arterialization on the morphology and function of the graft.

Heterotopik Yardımcı Sıçan Karaciğer Nakli ile Akut Karaciğer Yetmezliği Portal Ven arteriali Akış ayarlı

In acute hepatic failure auxiliary liver transplantation is an interesting alternative approach. The aim is to provide a temporary support until the ...

Tıp

Murine Heterotopic Heart Transplant Technique

It is now over forty years since this technique was first reported by Corry, Wynn and Russell. Although it took some years for other labs to become proficient in and utilize this technique, it is now widely used by many laboratories around the world. A significant refinement to the original technique was developed and reported in 2001 by Niimi. Described here are the techniques that have evolved over more than a decade in the hands of three surgeons (Plenter, Grazia, Pietra) in our center. These techniques are now being passed on to a younger generation of surgeons and researchers.
Based largely on the Niimi experience, the procedures used have evolved in the fine details - details which we will endeavor to relate here in such a way that others may be able to use this very useful model. Like Niimi, we have found that a video aid to learning is a priceless resource for the beginner.

The manuscript describes the steps required to perform the heterotopic heart transplant in the mouse.

Mürin Heterotopik Kalp Nakli Tekniği

It is now over forty years since this technique was first reported by Corry, Wynn and Russell. Although it took some years for other labs to become ...

An Immunological Model for Heterotopic Heart and Cardiac Muscle Cell Transplantation in Rats

Heterotopic heart transplantation in rats has been a commonly used model for diverse immunological studies for more than 50 years. Several modifications have been reported since the first description in 1964. After 30 years of performing heterotopic heart transplantation in rats, we have developed a simplified surgical approach, which can be easily taught and performed without further surgical training or background.
After dissection of the ascending aorta and the pulmonary artery and ligation of superior and inferior caval and pulmonary veins, the donor heart is harvested and subsequently perfused with ice-cold saline solution supplemented with heparin. After clamping and incising the recipient abdominal vessels, the donor ascending aorta and pulmonary artery are anastomosed to the recipient abdominal aorta and inferior vena cava, respectively, using continuous running sutures.
Depending on different donor-recipient combinations, this model allows analyses of either acute or chronic rejection of allografts. The immunological significance of this model is further enhanced by a novel approach of in-ear injection of vital cardiac muscle cells and subsequent analysis of draining cervical lymphatic tissue.

We describe a model of heterotopic abdominal heart transplantation in rats, implying modifications of current strategies, which lead to a simplified surgical approach. Additionally, we describe a novel rejection model by in-ear injection of vital cardiac muscle cells, allowing further transplant immunological analyses in rats.

Sıçanlarda Heterotopik Kalp ve Kardiyak Kas Hücre Nakli için İmmünolojik Model

Heterotopic heart transplantation in rats has been a commonly used model for diverse immunological studies for more than 50 years. Several modifications ...

İmmünoloji ve Enfeksiyon

Blood Circuit Reconstruction in an Abdominal Mouse Heart Transplantation Model

The surgical technique of heterotopic abdominal heart transplantation in mice is a standard model for research in transplantation immunology. Here, the established technique for a modified blood circuit reconstruction in a heterotopic abdominal heart transplantation model is presented. This method uses the intrathoracic inferior vena cava (IIVC) instead of the pulmonary artery of the donor heart for the anastomosis to the inferior vena cava of the recipient. It is facilitating and improving success rates for abdominal heart transplantation in mice.

A novel technique for blood circuit reconstruction in a heterotopic abdominal mouse heart transplantation model is demonstrated.

Abdominal Fare Kalp Nakli Modelinde Kan Devresi Rekonstrüksiyonu

The surgical technique of heterotopic abdominal heart transplantation in mice is a standard model for research in transplantation immunology. Here, the ...

A Simplified Model for Heterotopic Heart Valve Transplantation in Rodents

There is an urgent clinical need for heart valve replacements that can grow in children. Heart valve transplantation is proposed as a new type of transplant with the potential to deliver durable heart valves capable of somatic growth with no requirement for anticoagulation. However, the immunobiology of heart valve transplants remains unexplored, highlighting the need for animal models to study this new type of transplant. Previous rat models for heterotopic aortic valve transplantation into the abdominal aorta have been described, though they are technically challenging and costly. For addressing this challenge, a renal subcapsular transplant model was developed in rodents as a practical and more straightforward method for studying heart valve transplant immunobiology. In this model, a single aortic valve leaflet is harvested and inserted into the renal subcapsular space. The kidney is easily accessible, and the transplanted tissue is securely contained in a subcapsular space that is well vascularized and can accommodate a variety of tissue sizes. Furthermore, because a single rat can provide three donor aortic leaflets and a single kidney can provide multiple sites for transplanted tissue, fewer rats are required for a given study. Here, the transplantation technique is described, providing a significant step forward in studying the transplant immunology of heart valve transplantation.

This protocol describes a simple and efficient method for the transplantation of aortic valve leaflets under the renal capsule to allow for the study of alloreactivity of heart valves.

Kemirgenlerde Heterotopik Kalp Kapağı Transplantasyonu için Basitleştirilmiş Bir Model

There is an urgent clinical need for heart valve replacements that can grow in children. Heart valve transplantation is proposed as a new type of ...

A Modified Cuff Technique for Mouse Cervical Heterotopic Heart Transplantation Model

Cardiac allograft rejection limits the long-term survival of patients after heart transplantation. A mouse heart transplantation model is ideal for investigating the mechanism of cardiac allograft rejection in preclinical studies because of their high homology with human genes. This understanding would help develop unique approaches to improving patients&#39; long-term survival treated with cardiac allografts. In a mouse model, abdominal donor heart implantation is commonly performed with an end-to-end&#160;anastomosis to the recipient&#39;s aorta and inferior vena cava using stitches. In this model, the donor&#39;s heart is implanted by end-to-end anastomosis to the recipient&#39;s carotid artery and jugular vein by the modified-Cuff technique. The transplantation surgery is performed without stitching and thus may increase the survival of the recipient since there is no interference with the blood supply and venous reflux of the lower body. This mouse model would help investigate the mechanisms underlying the immunological and pathological (acute/chronic) rejection of cardiac allografts.

In the present protocol, a mouse heart transplantation model is used for investigating the mechanism of cardiac allograft rejection. In this heterotopic heart transplantation model, operation efficiency is improved, and the survival of cardiac grafts is ensured by a cervical end-to-end anastomosis of heart implantation using a modified Cuff technique.

Fare Servikal Heterotopik Kalp Transplantasyon Modeli için Modifiye Manşet Tekniği

Cardiac allograft rejection limits the long-term survival of patients after heart transplantation. A mouse heart transplantation model is ideal for ...

A Heterotopic Rat Heart Transplantation Model using Circulatory Death Donor Hearts

The objective of this protocol is to set up a rat heterotopic heart transplantation model with donation after circulatory death (DCD) donor hearts. There are two setups for this protocol: heart donor setup and recipient setup. In the heart donor setup, Sprague Dawley rats are anesthetized, endotracheally intubated, and ventilated. The right carotid artery is cannulated to deliver heparin and the paralytic agent vecuronium-bromide. The DCD process is initiated by terminating the ventilation. After 20 min, the heart is exposed and the aorta distal to the brachiocephalic branch is clamped. At 25 min from terminating the ventilator, ice-cold University of Wisconsin (UW) solution is perfused through the carotid catheter to flush the heart. The heart is procured by dividing the aorta, pulmonary artery, venae cavae, and pulmonary veins and stored in UW solution for implantation. In the recipient setup, the Lewis rat is anesthetized with isoflurane. Slow-release buprenorphine is administered subcutaneously to facilitate a smooth postoperative recovery. Through a midline abdominal incision, the infra-renal aorta and the inferior vena cava are isolated and clamped with an atraumatic vascular clamp. The donor heart aorta and pulmonary artery are sutured to the recipient abdominal aorta and vena cava, respectively, with a running 8-0 Prolene. The vascular clamp is removed to reperfuse the heart. The abdominal wall is closed and the rat is recovered. After a set interval (24 h to 2 weeks), the recipient rat is anesthetized, the transplanted heart is exposed, and a balloon-tip-catheter is inserted into the left ventricle via the apex to record developed pressure and dP/dt using a data acquisition system. The heart tissue is collected for histology, immunology, or molecular analysis. A successful DCD donor rat heart transplantation model will allow further studies on the cardioprotective approaches to improve heart transplantation outcomes from DCD donors.

Here, we describe the preparation and technical details of murine heterotopic heart transplantation utilizing a circulatory death donor heart.

The objective of this protocol is to set up a rat heterotopic heart transplantation model with donation after circulatory death (DCD) donor hearts. There ...

Rat Model of Normothermic Ex-Situ Perfused Heterotopic Heart Transplantation

Heart transplantation is the most effective therapy for end-stage heart failure. Despite the improvements in therapeutic approaches and interventions, the number of heart failure patients waiting for transplantation is still increasing. The normothermic ex situ preservation technique has been established as a comparable method to the conventional static cold storage technique. The main advantage of this technique is that donor hearts can be preserved for up to 12 h in a physiologic condition. Moreover, this technique allows resuscitation of the donor hearts after circulatory death and applies required pharmacologic interventions to improve donor function after implantation. Numerous animal models have been established to improve normothermic ex situ preservation techniques and eliminate preservation-related complications. Although large animal models are easy to handle compared to small animal models, it is costly and challenging. We present a rat model of normothermic ex situ donor heart preservation followed by heterotopic abdominal transplantation. This model is relatively cheap and can be accomplished by a single experimenter.

Here, we present an assessment protocol of a heterotopically implanted heart after normothermic ex situ preservation in the rat model.

Normotermik Ex-Situ Perfüze Heterotopik Kalp Transplantasyonunun Sıçan Modeli

Heart transplantation is the most effective therapy for end-stage heart failure. Despite the improvements in therapeutic approaches and interventions, the ...

Modified Heterotopic Abdominal Heart Transplantation and a Novel Aortic Regurgitation Model in Rats

Over the past 50 years, many researchers have reported heterotopic abdominal heart transplantation in mice and rats, with some variations in the surgical technique. Modifying the transplantation procedure to strengthen the myocardial protection could prolong the ischemia time while preserving the donor&#39;s cardiac function. This technique&#39;s key points are as follows: transecting the donor&#39;s abdominal aorta before harvesting to unload the donor&#39;s heart; perfusing the donor&#39;s coronary arteries with a cold cardioplegic solution; and topical cooling of the donor&#39;s heart during the anastomosis procedure. Consequently, since this procedure prolongs the acceptable ischemia time, beginners can easily perform it and achieve a high success rate.
Moreover, a new aortic regurgitation (AR) model was established in this work using a technique different from the existing one,&#160;which is created by inserting a catheter from the right carotid artery and puncturing the native aortic valve under continuous echocardiographic guidance. A heterotopic abdominal heart transplantation was performed using the novel AR model. In the protocol, after the donor&#39;s heart is harvested, a stiff guidewire is inserted into the donor&#39;s brachiocephalic artery and advanced toward the aortic root. The aortic valve is punctured by pushing the guidewire further even after the resistance is felt, thus inducing AR. It is easier to damage the aortic valve using this method than with the procedure described in the conventional AR model. Additionally, this novel AR model does not contribute to the recipient&#39;s circulation; therefore, this method is expected to produce a more severe AR model than the conventional procedure.

This study demonstrates a reproducible heterotopic abdominal heart transplantation technique in rats that beginners can learn and perform. Additionally, a novel aortic regurgitation model in rats is generated by performing heterotopic abdominal heart transplantation and damaging the donor's aortic valve using a guidewire after harvesting.

Modifiye Heterotopik Abdominal Kalp Transplantasyonu ve Sıçanlarda Yeni Bir Aort Yetersizliği Modeli

Over the past 50 years, many researchers have reported heterotopic abdominal heart transplantation in mice and rats, with some variations in the surgical ...